Affiliation:
1. Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts
Abstract
ABSTRACT
Pseudomonas aeruginosa
uses a dedicated type III secretion system to deliver toxins directly into the cytoplasm of host cells. While progress has been made in elucidating the function of type III-secreted toxins in vitro, the in vivo functions of the type III-secreted exoenzymes are less well understood, particularly for the sequenced strain PAO1. Therefore, we have systematically deleted the genes for the three known type III effector molecules (
exoS
,
exoT
, and
exoY
) in
P. aeruginosa
PAO1 and assayed the effect of the deletions, both singly and in combination, on cytotoxicity in vitro and in vivo. We found that the type III secretion system acts differently on different cell types, causing an
exoST
-dependent rounding of a lung epithelial-like cell line in contrast to causing an
exoSTY
-independent but translocase (
popB
)-dependent lysis of a macrophage cell line. We utilized an in vivo competitive infection model to test each of our mutants, examining replication in the lung and spread to secondary sites such as the blood and spleen. Type III mutants inoculated intranasally exhibited only a minor defect in replication and survival in the lung, but
popB
and
exoSTY
triple mutants were profoundly defective in their ability to spread systemically. Intravenous injection of the mutants indicated that the type III secretion machinery is required for survival in the blood. Furthermore, our findings suggest that the effector-independent
popB
-dependent cytotoxicity that we and others have observed in vitro in macrophage cell lines may not be of great importance in vivo.
Publisher
American Society for Microbiology
Subject
Infectious Diseases,Immunology,Microbiology,Parasitology
Cited by
160 articles.
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